| Studies have shown that the reproductive activity of animals by the hypothalamus- pituitary- gonadal axis regulation, basophils its anterior pituitary gonadotropin synthesis and secretion of mammalian sexual development, sexual maturity, physical characteristics of the dominant cycle plays an important factor. Wherein Follicle stimulating hormone and luteinizing hormone directly involved in the regulation of dams oestrus, ovulation and other reproductive process. Thus, form and expression of Tibetan sheep pituitary FSH and LH cells, Tibetan sheep reproductive activities have importance value.To studied the role of changes in the expression in the pituitary FSH and LH cells and the secretion of hormones in the reproductive activities of Ganjia Tibetan sheep estrous cycle. This study(1) the changes follicle-stimulating hormone and luteinizing hormone cells pituitary distribution of m RNA expression were studied on different Ganjia Tibetan sheep estrous cycle;(2)the dynamic FSH(FSH) and luteinizing hormone(LH) secretion variation were studied in estrous cycle at different stages of pituitary, and observed anestrus pituitary FSH and LH change. This study was select 2.5-3.5 years old, the 30 sheep of estrous cycle and anestrus were killed for pituitary and plasma that they were healthy nonpregnant female. according to the different stages of the estrous cycle is divided into four groups: Proestrus, Oestrus, Metestrus, Dioestrum. 2.5-4 years of age, The estrous cycle and anestrus were killed for pituitary and plasma that they were healthy nonpregnant female sheep 8. Applications(1)HE staining cell morphology and distribution characteristics of each group Ganjia Tibetan sheep pituitary;(2) immunohistochemistry SP method was for each group FSH and LH cells reservoir distribution expression sheep pituitary tissue were studied, and the optical density expression image pro-plus image analysis pituitary tissue FSH and LH immunoreactive product;(3) real-time quantitative PCR( RT-PCR) technology, FSHβ and LHβm RNA of pituitary at different stages of the estrous cycle Ganjia Tibetan sheep were expressed.(4) enzyme-linked immunosorbent assay(ELISA) was measured in each group and analysis FSH and LH content of pituitary the dynamic changes of the estrous cycle Tibetan sheep.The results are as follows:1.HE staining showed: eosinophils larger, round or oval, eosinophilic cytoplasm coloration, nuclear round, colored deep within the pituitary of Ganjia Tibetan sheep estrous cycle; basophils volume maximum, circular or polygonal, cytoplasmic basophilic coloration, nuclear round, lighter color; chromophobe smallest cytoplasm slightly stained, ill-defined cell nuclei round or irregular in shape, lighter color of. And three kinds of physiological activities of the cells have been changed in the estrous cycle. Eosinophils in the estrous cycle and anestrus secretory granules and cytoplasmic coloration stable; basophils in proestrus, estrus, late estrus cytoplasmic secretory granules and gradually increased the number of coloring in diestrus and anestrus reduced: lightly stained chromophobes little cytoplasmic granules, but the number of changes in the estrous cycle is obvious, and in larger quantities during estrus and anestrus.2.Immunohistochemistry showed:FSH and LH immunoreactive cells were mainly distributed in the distal portion of Ganjia Tibetan sheep estrous cycle, an intermediate portion of the pituitary little; less estrus FSH and LH positive cells, irregular shape, lightly stained cytoplasm; the number of post-estrus and diestrus increased, cells and medium-sized, oval or polygonal shape, cytoplasmic stain, and diestrus LH positive cells reached maximum; increase the maximum number of FSH proestrus cells large, round or oval, cytoplasm stain; FSH than LH cell positive cytoplasmic stain. FSH and LH percentage of positive cells in the pituitary number(oestrus,metestrus,dioestrum, proestrus): FSH was 9.78%, 11.37%, 12.81%, 13.05%,; LH 8.72% 12.73%, 14.25%, 13.72%, and the difference between each period were not significant(p> 0.05).3.IPP estimated average optical density image display: FSH expression of oestrus in lower increase in metestrus, dioestrum and proestrus maximized while other times significantly different(P <0.05); LH expression in oestrus low period, late estrus, diestrus and proestrus were increased and the difference was not significant(p>0.05). LH expression except in oestrus FSH lower than, other periods were higher than FSH.4.Real-time quantitative PCR showed: FSHβ and LHβ m RNA at different stages of the estrous cycle Ganjia Tibetan sheep pituitary were expressed. FSHβ relative expression(proestrus, oestrus,metestrus,dioestrum) was 0.2359 ± 0.017,1 ± 0,3.5475 ± 0.8017,3.6667 ± 1.4492, made proestrus and oestrus, metestrus and dioestrum difference highly significant(p<0.01), oestrus and metestrus and dioestrum difference estrus was significantly(p<0.05), metestrus and metestrus was no significant difference(p>0.05); LHβ relative expression was 0.0259 ± 0.0038,1 ± 0,16.1091 ± 3.2674,0.5638±0.1962, proestrus and oestrus, metestrus and dioestrum differ significantly(p<0.01), oestrus and metestrus difference was significant(p<0.01), and dioestrum the difference was not significant(p>0.05), metestrus and dioestrum was significantly different(p<0.01).5.The ELISA test showed that: FSH concentrations during pituitary dioestrum maximum(8.228 m IU / ml) of Ganjia Tibetan sheep estrous cycle, and other times the difference was significant(P <0.05); pituitary LH concentration in dioestrum maximum(10.629 m IU / ml), each time the difference was not significant(P>0.05);... |
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